Kinetic and Physical Studies of Cell Death Induced By Chemotherapeutic Agents Or Hyperthermia

The kinetics of three physical parameters: cell density, relative cytoplasmic viscosity and DNA stability to denaturation have been measured during the period preceding cell death induced by hyperthermia, methylprednisolone and a series of cancer chemotherapeutic agents. This series of measurements employed cultured human lymphoblastoid cells as an experimental system to establish the changes that can be observed in the early stages of cell death, prior to applying such measurements to tissue biopsies from solid human tumours. Cell death, induced by hyperthermia up to 43°C, methylprednisolone, vincristine, 5-fluorouracil, BCNU and melphalan, showed essentially identical and reproducible changes corresponding to those which characterize programmed cell death (apoptosis). Such changes could also be observed following hyperthermia above 43°C, but reproducibility was poor and increasing damage to the cell membranes was evident. In cells treated with adriamycin or methotrexate, cell sub-populations showing an increase in cell density were not detected. Measurements of DNA stability were readily performed by flow cytofluorometry thus allowing rapid quantitation of the fraction of cells in the early stages of cell death. Modified flow cytometric instrumentation would further allow measurement of cytoplastic viscosity as an additional parameter to indicate entry into programmed cell death. This suggests that these measurements could readily be applied to cell suspensions derived from tumour tissue biopsies for a more accurate assessment of tumour growth rate, and to allow monitoring of response to therapy in sequential tumour biopsies.     

Association of genetic polymorphisms with embryonic mortality in the chicken

Summary Association of the egg-white genotypes of dams with the mortality of their embryos was studied in 1966 in the pure and crossbred embryos of dams of two relatively noninbred Light Sussex substrains, 6D and 6F. These had been derived two generations earlier, in 1964, by equal division of strain 6, which we previously studied in 1962; while the sires of the crosses came from two Rhode Island Red/New Hampshire substrains, 5D and 5F, similarly derived. The effects were estimated parametrically for the three loci Tf, II and III singly and in two- and three-way interaction.In the first week of incubation, Tf had an additive effect in all matings, while II (and perhaps III) had a near-additive effect in the pure matings only. There was an additive X additive interaction effect of II X III in the last six days of incubation, which was consistent over all matings except 5F X 6F. The only effect to influence the total mortality over the whole of incubation was the three-way additive X additive X additive interaction, and there was substantial agreement in this over the matings. Some triple homozygotes were superior and others inferior to the genotypes with one or more loci heterozygous.This evidence of multiple interaction between loci affecting fitness is discussed in relation to hypotheses for the maintenance of genetic variability in populations.     

An estimation of tissue damage and thermal history in the cryolesion

The cooling and thawing rates at fixed sites within an expanding ice ball have been recorded using a temperature probe containing six integral thermocouples. The cell changes relevant to these sites have been followed through the 48 hr postfreeze-thaw period by light and electron microscopy. The results suggest that cooling rates that are compatible with cell survival in cell suspension are associated with uniform tissue cellular death in an organized tissue.     

Studies on brain-cortex slices: Differences in the oxidation of 14C-labelled glucose and pyruvate revealed by the action of triethyltin and other toxic agents

1. The rate of appearance of ¹⁴CO₂ from [6-¹⁴C]glucose and [3-¹⁴C]pyruvate was measured. Pyruvate is oxidized to carbon dioxide twice as fast as glucose, although the oxygen uptake is almost the same with each substrate. 2. The presence of 30μm-2,4-dinitrophenol increases the output of ¹⁴CO₂ from [6-¹⁴C]glucose sixfold whereas the oxygen uptake is not quite doubled. Similar results are obtained with 0·1m-potassium chloride. The stimulating action of these two agents on the output of ¹⁴CO₂ from [3-¹⁴C]pyruvate is much less than on that from [6-¹⁴C]glucose. 3. The effects of oligomycin, ouabain and triethyltin on the respiration of control and stimulated brain-cortex slices were studied. Triethyltin (1·3μm) inhibited the oxidation of [6-¹⁴C]glucose more than 70%, but did not inhibit the oxidation of[3-¹⁴C]pyruvate. [3-¹⁴C]pyruvate. 4. The production of lactic acid by brain-cortex slices incubated with glucose is twice as great as that with pyruvate. Lactic acid increases two and a half times in the presence of either triethyltin or oligomycin when the substrate is glucose, but is no different from the control when the substrate is pyruvate. 5. With kidney slices the production of lactic acid from glucose is very low. It is increased by oligomycin but not by triethyltin. 6. The results are discussed in terms of the oxidation of the extramitochondrial NADH₂ produced during glycolysis.     

Medicolegal considerations in the initiation and termination of resuscitation in Canada.

Medicolegal issues in cardiopulmonary resuscitation (CPR) and emergency cardiac care were considered in the United States by the National Conference on Cardiopulmonary Resuscitation in 1985. This paper discusses these issues in the Canadian context. Although there is little legislation or case precedent in Canada to guide providers of CPR in decision-making, there appears to be little risk of liability or prosecution for competently rendered care. Providers should be cautious in withholding or withdrawing resuscitative measures from incompetent patients when brain death has not occurred and cardiovascular unresponsiveness has not been demonstrated. However, resuscitation may be withheld when a competent patient refuses it or if there is another medically and legally valid reason to do so.     

Flow cytometric detection of ornithine decarboxylase activity in epidermal cell subpopulations

Using flow cytometry in combination with membrane permeabilization techniques to enhance binding of antibodies with immunoreactive protein within the cytoplasm, we have developed a method to examine the ornithine decarboxylase (ODC) activity present within subpopulations of epidermal cells following acute and chronic exposure to the phorbol ester tumor promoter 12-0-tetradecanoylphorbol-13-acetate (TPA). The method described has the sensitivity to detect basal levels of ODC as well as increases in ODC at early time points following treatment with TPA and has the additional advantage of allowing subpopulation identification and characterization.